To achieve the catalytic hydrolysis of phosphodiesters, nature often capitalizes upon a multifunctional approach that involves cooperativity between three catalytic groups: metals, guanidiniums, and general bases. Despite the abundant natural examples, human kind has not yet mastered the design principles for constructing artificial enzymes that display such functional group cooperativity. The immediate goal of the proposed research is to delineate these design principles in the construction of a metallonuclease. The three catalytic groups common to nucleases will be systematically combined in artificial enzymes, and the energetics of catalysis will be studied. By examining the energetics of functional group cooperativity, we seek information that will be essential to the design of efficient artificial metallonucleases. Three Programs involving incremental increases in complexity are proposed: Bi, Tri, and Tetrafunctional Catalysis. As the mechanisms of catalysis in each Program are deciphered, the complexity will increase via the addition of more functional group. Deciphering each mechanism will consist of thermodynamic and kinetic studies of several structurally related synthetic receptors. For example, in each Program the angles and distances between the functional groups will be varied as a means of probing their preorganization and complementarity for phosphorane-like transition states. In addition, each Program will also involve an energetic analysis derived from Transition State Theory (TST). The TST analysis allows for the separation of electrostatics and shape changes in the stabilization of phosphorane-like transition states. The focus of all three studies will be upon delineating those factors that allow confident construction of multifunctional metallonucleases. Understanding the physical organic chemistry of catalysis will lead to both a better understanding of enzymology, and the production of small molecules with catalytic activity for applications as pharmaceuticals. Both of these aspirations can only be achieved after many generations of synthetic catalysts have been produced, and the rules for combining functional groups have been deciphered. Achieving these rules, and thereby the ability to rationally produce catalysts for biomedical applications, is the long term goal of the proposed research.